Project Summary Project 1 ? Post-transcriptional mechanisms of muscle atrophy prevention in hibernating mammals. Reduced skeletal loading leads to muscle atrophy in humans and most mammals. Disuse muscle atrophy represents a significant clinical problem for patients during prolonged periods of immobilization and bed rest. Bears and ground squirrels are largely inactive during hibernation, but they show less muscle loss than would be anticipated over such a prolonged period of physical inactivity. This suggests that hibernating mammals have unique natural adaptation to musculoskeletal disuse. Although attenuation of muscle atrophy in hibernating bears and ground squirrels is well documented, molecular mechanisms underlying this important adaptation are not known. The proposed research focuses on post-transcriptional regulation in muscle of hibernating mammals that includes differential microRNAs expression and regulation of protein synthesis. Our goal is to identify, through microRNA expression and ribosome profiling, candidate microRNAs and their target transcripts, transcripts translated to proteins, and metabolic and signaling pathways that underlie the ability to reduce disuse muscle loss during hibernation. To achieve this goal we will accomplish the following Specific Aims (SA). SA1. Analysis of microRNA expression profiles across different states of the hibernation cycle in muscle of the black bear and arctic ground squirrel. SA2. Gene prediction and annotation of de novo genome assembly of the arctic ground squirrel. SA3. Analysis of gene expression at the protein level using ribosome profiling across different states of the hibernation cycle in muscle of the arctic ground squirrel. Once completed, genome-wide microRNA and ribosome profiling will detect sufficiently large number of differentially expressed genes for comprehensive pathway analysis elucidating the functional significance of transcriptional changes. We expect comparison of functional gene groups and pathways enriched by co-regulated genes between two evolutionary distant species with different hibernation modes to reveal a common molecular program and druggable targets for future study and development of improved treatments for and prevention of disuse muscle atrophy.